Blood clotting Factor VIII is a trace plasma glycoprotein that is found in mammals and is involved as a cofactor of IXa in the activation of Factor X. An inherited deficiency of Factor VIII results in the bleeding disorder haemophilia A, which can be treated successfully with purified Factor VIII. The Factor VIII can be extracted from blood plasma or can be produced by recombinant-DNA-based techniques. In the plasma, it circulates as a complex with von Willebrand Factor (vWF).
Recombinant Factor VIII (rFVIII) can be produced by Chinese Hamster Ovary (CHO) cells transfected with a vector carrying a DNA sequence encoding the Factor VIII molecule. In some cases, recombinant Factor VIII is co-produced with recombinant von Willebrand Factor (rvWF), which stabilises the Factor VIII. Such co-production can involve the co-culturing of respective cell lines that express FVIII and vWF, or the co-expression of the two proteins in the same cell. See U.S. Pat. No. 5,250,421 (Genetics Institute) and Kaufman et al (1989) Mol. Cell. Biol. 9, 1233-1242.
In a typical process for preparing recombinant Factor VIII, cells are cultured in a medium and secrete Factor VIII into the medium. Factor FVIII may then be purified from the medium, optionally as a complex with vWF.
Recombinant Factor VIII is expensive to produce due to the relatively low yields obtained in processes known in the art. The yield per cell tends to be low compared to the yield that might be obtained for other recombinant proteins. If the culture medium is not supplemented with animal products, such as serum, the medium may support only relatively low cell densities. This reduces the yield per volume of medium. However, it is desirable not to supplement the culture medium with animal products in order to reduce the risk of contamination with viruses and other transmissible agents. Animal-protein-free media for the production of FVIII are known from U.S. Pat. No. 6,936,441 (Baxter AG), for example.
The present invention provides processes for producing blood proteins, including rFVIII, in which the yield is improved compared to processes known in the art.